Background: (3±1°C, 85±5% RH). Results: The results of

Background: Fresh pistachio can be affected by physiological and biochemical
changes both during harvest and after harvest, and its optimum duration of
storage cannot therefore be prolonged.

Materials and
methods: In order to maintain the quality of
fresh pistachio fruits (Pistacia vera L. Ahmad Aghaei),
carboxymethyl cellulose at different concentrations accompanied by different
concentrations of Zataria multiflora essential oil was used to produce
an edible coating. The effects of these combinations were examined on the
quality, shelf life, and sensory properties of the fresh fruit during a storage
period of 32 days (3±1°C, 85±5% RH).

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Results: The results of sensory and instrumental tests during storage on
days 8, 16, 24, and 32 showed that coated samples with 1.5% (w/v) carboxymethyl
cellulose used in combination with Zataria multiflora essential oil had
the longest shelf life compared to the other treatments. Among the treatments
containing 1.5% (w/v) carboxymethyl cellulose, the lowest weight loss and the
highest kernel carbohydrate, total soluble solid, phenolic compounds, and
antioxidant activity were observed in the treatments containing 1.5% (w/v) carboxymethyl
cellulose accompanied by 0.4% (w/v) and 0.2% (w/v) Zataria multiflora
essential oil.

Conclusion: Application of 1.5% (w/v) carboxymethyl cellulose in combination
with 0.4% (w/v) and 0.2% (w/v) Zataria multiflora essential oil
effectively maintained the quality of fresh pistachio during the 32 days of storage.


Keywords: Fresh pistachio; edible coating; carboxy methyl cellulose; essential
oil; Zataria multiflora




1. Introduction

Pistachio is an agricultural product with high nutritional value. This
fruit is a good source of vitamins A, B1, B2, B6,
and E, and minerals such as iron, phosphorus, selenium, zinc, and fatty acids.
Fatty acids are precursors in the synthesis of prostaglandins and ultimately
prevent the accumulation of erythrocytes in the blood (1).

Recently, the application of natural and biodegradable coatings instead of artificial waxes has increased (2). Edible coatings consist of a thin layer of nutrients
that form directly on the fruit’s surface. These coatings have the potential to
create a selective barrier to moisture, CO2, and oxygen. Edible coatings may
control the internal atmosphere of the fruit, minimize the respiratory rate of
the product, and delay the evaporation of water and moisture loss (3). Edible coatings increase the shelf life of fresh fruits and vegetables by
reducing metabolic processes and microbial growth (4).  

Cellulose derivatives are linear polysaccharides composed of ?-1 and 4 glucose
units with methyl, hydroxypropyl, or carboxyl substitutions. Carboxymethyl
cellulose is one of the most common cellulose derivatives used in the
preparation of edible films (5). It is linear, has a long chain, and is a
water-soluble, anionic, and non-allergenic polysaccharide (5). This coating, in
combination with antioxidant and antimicrobial agents, can effectively prevent
the growth of fungi and microorganisms (5). The respiration rate is
one of the major factors contributing to the postharvest loss of fruit (6). Azarakhsh et al. showed that the respiration
rate of fresh-cut pineapple samples coated with an edible alginate coating was significantly
lower than the uncoated samples during storage (6). This reduced respiration rate is achieved by
the coating creating an internal modified atmosphere which reduces the exchange
of carbon dioxide and oxygen between the environment and the coated fruit (7).
Also, Asgar at al.
observed that papayas coated with chitosan showed a lower respiration rate and
ethylene production during storage (8). Reductions in respiration rate and
ethylene production as a result of edible coating has also been reported by
many researchers reporting research on various fruits, such as grapes,
strawberries, papayas, tomatoes, and mangos (9). The application of this cellulose coating on pecan
can limit the contact of oxygen with kernels and 
the exchange of gases by acting as a barrier. This results in a reduced
oxidation of lipids in the kernel (10). Edible coatings have the potential to carry
active ingredients, such as anti-browning agents, coloring products,
antimicrobials, and essential oils (11). The results of Raeisi et al.’s study showed that edible carboxymethyl cellulose coating
with the essential oil of Zataria
multiflora and grape seed extract is very effective in reducing the undesirable
chemical reactions in fish meat during storage (11). According to the research conducted by Dhall Group,
edible coatings should basically be resistant to water. They should not show
destructive behaviors on O2 or cause the accumulation of excessive
amounts of CO2 (2). Edible coatings should have minimum permeability
to water vapor, and their presence is expected to improve the fruit appearance
and gloss, while preventing the fruit from becoming sticky. Edible coatings
should be economical and optically transparent during the time of storage.

Therefore, this study aimed to evaluate the effects of different
concentrations of carboxymethyl
cellulose (CMC) edible coating in combination with different concentrations of Zataria multiflora essential
oil on weight loss, total soluble solids (TSS), carbohydrate, phenolics,
antioxidant activity, and sensory properties of fresh pistachio during 32 days
of storage.

2. Materials and Methods

2.1 Materials

2.1.1. Fruit

This study was conducted on an important commercial pistachio cultivar
called Ahmad Aghaei. The samples were prepared from a pistachio
orchard located in Pistachio Research Center in the city of Rafsanjan.


2.1.2. Coatings

Carboxymethyl cellulose as an edible coating
and glycerol as a plasticizer in the edible coating were supplied from Sigma-Aldrich
(Steinheim, Germany). The Essential oil of Zataria multiflora, was purchased from Barij
Essence (Esfahan, Iran).


2.1.3. Preparation of samples and treatments

The fresh Ahmad
Aghaei pistachios were harvested at maturity and transferred to the laboratory,
and pistachio fruits werethen isolated from the cluster in order to treat.

The coating solution was prepared following the
method used in Arnon et al. (12). Briefly, a solution of an appropriate amount
of carboxymethyl cellulose powder in distilled water was prepared by heating at
80°C and stirring to form a clear solution. Then, 1% (w/v) glycerol was added
to the solution as a plasticizer. Different concentrations of Zataria multiflora essential oil (0.0, 0.2, and 0.4% (w/v)) were added to
the mixture. Finally, all formulations were homogenized for 10 min.



2.1.4. Treating and storage of fruits

Fresh Pistachios were immersed in coating solutions for 3
min and were then air-dried at room temperature for 1 min. They were then sorted
to 200-gr packs in each polypropylene dish and stored at 3±1°C and 85±5% RH for
32 days. Fruits without coating and essential oil were also placed at the same
condition as control. Weight loss, total soluble solids,
carbohydrate, phenol, antioxidant activity, and hedonic test were evaluated after 0, 8, 16, 24, and 32 days of storage.


2.2. Parameters assay

2.2.1. Weight loss

Fruit weight loss percentage was measured using the
following equation (Eq. 1):

                      (Eq. 1)


2.2.2. Total phenolic compounds and total antioxidant activity

To estimate the phenolic value of the pistachio, the method reported by Singleton
et al. (13) was used. Briefly, 0.5 gr of the
kernel was homogenized with 3 mL of 85% (v/v) methanol, and the resulting
mixture was then centrifuged at 10,000 rpm for 15 min. Then, 150 µL of the
supernatant was transferred to a test tube where 75 µL of Folin-Ciocalteu
phenol reagent was added. After an incubation period of 5 min, 600 µl of 7% (w/v)
Na2CO3 was added to the solution, which was then mixed
well and kept in dark for 1.5 h. The samples were vortexed. Their absorbance
was then measured at 760 nm.

Antioxidant activities were measured using the method presented by the Brand-Williams Group (14). To do this, an extraction procedure similar
to the one applied for phenolic value was conducted. After the extraction, 250
?l of the supernatant was mixed with 250 ?l of distilled water and
centrifuged for 10 min. Then, 75 ?l of the combined solution was mixed with
2.925 ml DPPH 85% (w/v), and the absorbance of each solution was recorded at
517 nm. After incubation in dark for 30 min, the absorbance was again measured
at the same wavelength.



2.2.3. Total soluble solids (TSS) and carbohydrate

The TSS value
was measured as an average of 10 fruits in each replicate and assessed by a
digital refractometer (ATAGO,PAL-1model Japan). To determine carbohydrate content, 0.5 g of pistachio ash (without oil) was
homogenized with 5 ml of 95% (v/v) ethanol. The extraction was repeated twice using 5
ml of 70% (v/v) ethanol, and the obtained
mixture was then centrifuged at 3,500 rpm for 20 min. After that, 100 ?l of the
alcoholic extract was mixed with 3 ml of freshly prepared anthrone (150 mg of
anthrone in 100 ml of sulfuric acid 72% (v/v)). The solution was placed in a
water bath at 90°C for 10 min. Then, absorption rate was measured at 625 nm (15).



2.2.4. Sensory analysis

For sensory analysis, eight trained panelists were selected from the Pistachio Research
Center staff. The sensory evaluation form had a score scale from 0 to 15, the
lowest to the highest admission. The panelists assessed the parameters hull color, pistachio
color, taste, flavor, strange flavor and odor, juiciness texture, and overall
visual and flavor acceptance based on the evaluation forms.


Statistical analysis

This experiment
was performed with three factors in a factorial design based on a completely
random design. Each value is the average of three replications. Sources of
variation were CMC edible coating, Zataria multiflora essential oil, and the duration of storage. The experimental data were subjected to analysis of variance
(ANOVA) by using the SAS 9.1 statistical software.


3. Results

3.1. Weight loss

As depicted in Fig.1, the weight loss of all coated fruits increased after
32 days of storage. The highest percentages of weight loss were observed in the
control (8.28%) and the fruits coated by 0.2% (w/v) Zataria mulrifolra essential oil (9.06%) and the lowest was associated with
1.5% (w/v) CMC. In 1.5% (w/v) CMC edible coating fruits, the weight loss
gradually decreased with increasing the Zataria mulrifolra essential oil concentration, and finally 1.5% (w/v) CMC treatment in
combination with 0.4% (w/v) Zataria mulrifolra essential oil showed the
least weight loss (4.43%) after 32 days of storage.


3.2. TSS and carbohydrate content

The highest and the lowest TSS% were observed in the control and the fruits
treated with 1.5% (w/v) CMC, respectively (Fig. 2A). In the present study, the
coatings enriched with 0.2% (w/v) and 0.4% (w/v) Zataria mulrifolra essential oils were more effective in reducing
metabolic processes in fresh pistachio compared with the other treatments.

The results indicated that there were significant differences in the
carbohydrate content between uncoated and coated fruits during the storage
period (Fig. 2B). It was found that treatment with 1.5% (w/v) CMC enriched with
0.2% (w/v) and 0.4% (w/v) Zataria mulrifolra essential oil showed the
highest carbohydrate content in comparison with the other treatments in 32 days
of storage.


3.3. Phenolic and Antioxidant activity

As shown in Fig. 3A, the lowest and the highest contents of phenolics were observed
in the control (140.15 mg gallic acid per 100 g fresh weight) and the fruits coated
with 1.5% (w/v) CMC. Of the coatings containing 1.5% (w/v) CMC, the one enriched
with 0.4% (w/v) Zataria mulrifolra essential oil showed the highest phenolics contents (221.59 mg gallic acid
per 100 g fresh weight), which was found to be statistically significant.

Antioxidant activity of the kernel was significantly affected by the
treatment composition as well during storage (Fig.3B). During the storage period,
the fruits treated with the coating containing 1.5% (w/v) CMC and 0.4% (w/v)
essential oil showed the highest antioxidant activity in comparison to the other
treatments. For example, on day 32, the fruits coated with 1.5% (w/v) CMC accompanied
by 0.4% (w/v) essential oil showed 24.79 % antioxidant activity, while the
control sample showed 14.66 %.


3.4. Sensory

evaluation was conducted based on hull color, taste, odor, strange flavor and
odor, juiciness texture, and overall visual and flavor acceptance for both
coated and uncoated samples during the storage period of 32 days at 3±1°C (Fig.
4). The incorporation of 0.2% (w/v) and 0.4% (w/v) zataria mulrifolra
essential oil into the CMC-based coating formulation had desirable effects on the
sensory attributes of the coated fruits. Of all the coatings, the one enriched
with 0.4% (w/v) zataria mulrifolra showed more positive effects
on the overall acceptability of samples (11.41).


4. Discussion

In 2012, the Wittaya Group reported that edible coatings containing
plasticizers are good inhibitors for moisture loss, and they can specifically
reduce the rate of moisture loss from seeds (16). The weight loss of fresh
pistachios coated by the CMC in our study was consistent with the results
reported by the Albanese Group in 2007 (17), who demonstrated that the
immersion of citrus fruits in CMC coating effectively reduced weight loss. It
was suggested that the positive effect of this coating was capable of reducing respiration
rate due to the development of an amorphous glass on the citrus surface which held
back water evaporation. In 2007, the Lins Group stated that edible coatings
provided effective barriers against oxygen, carbon dioxide, and water vapor
transmissions and, as a result, they could reduce moisture loss (18).  Also, in 2013, the Athmaselvi Group reported that
tomatoes coated with Aloe vera showed a gradual decrease in the TSS
during storage (19), a result in line with the findings of our study. This may
be due to the break-up of pectin and carbohydrates, partial hydrolysis of
protein, and decomposition of glycosides into subunits during respiration,
which causes a decrease in the total soluble solids (19). Furthermore, the Athmaselvi Group reported
that sugar content in coated tomatoes was higher compared to the corresponding
control, due to the controlled atmosphere around the fruit created by the
edible coating that in turn was obtained by reducing respiration and
transpiration loss (19).

Phenolic compounds are natural antioxidants that can be
found in different vegetable sources (20). The relationship between the amount
of the total phenolics and the antioxidant properties has been studied in many
fruits and vegetables (20). It is believed that phenolic components are
considerably involved in the antioxidant capacity (20). The Guerreiro Group
previously reported that the antioxidant capacity of Arbutus unedo fresh
fruits increased when the fruits were treated with alginate edible coatings
enriched with eugenol and citral essential oils (4), which is in line with our
results. The decrease in antioxidant activity during storage may be attributed
to the destruction of cell structures as the fruit undergoes senescence (21).
Edible coatings may provide a barrier to reduce oxygen consumption and
therefore reduce oxidative processes (22). The antioxidant effects of some
essential oil compounds may also contribute to the maintenance of antioxidant
activity in fruits (23). Furthermore, in 2008, the Oms-Oliu Group found that using
edible coatings was effective in reducing total phenolic compounds and
antioxidant capacity in fresh-cut melon during storage (24). In 2015, the Dashipour
Group reported that the highest total phenolic and antioxidant activity was
observed in CMC films incorporated with Zataria multiflora essential oil
(5). In our study, the increase in antioxidant activity was related to the
enhancement of phenolics in the pistachio kernel. Also, some researchers have
stated that the accumulation of phenolic compounds can be a result of an increase
in antioxidant activity (24-25).

The addition of essential oils to edible coatings instead of the direct application
of the essential oils can cause the oils to be released gradually. This
improves their performance and efficiency. Our results are in line with those previously
reported by the Khoshnodinia Group in 2013 who showed that the application of
antioxidant-gelatin combined with an edible coating on pistachio had greater
effects on the sensory scores in comparison with the absence of the antioxidant
(26). Since edible coatings are usually eaten along with the fruits, investigation
of their sensory properties is of great importance (27). The sensory results
obtained in the present study are also in harmony with those reported by the Arnon
and the Tzoumaki groups (12, 28). In 2006, the Baldwin Group indicated that CMC
can influence the sensory parameters related to the pecan kernel (10). CMC-containing
coatings also improve the appearance and gloss of the coated fruit, maintain the
fruit quality and flavors, and fully covers the fruit surface (28). Moreover, this
coating is deemed economically affordable and is considered a homogeneous and
transparent material. Also, CMC-based coatings possess good stability and can
be easily prepared (28).

Thymol and carvacrol are the most important and abundant components in thyme
essential oil. These components belong to the group of phenols and have
antifungal properties (29). It has been also reported that the phenolic
compounds available in thyme have inhibitory effects on the growth of Aspergillus
flavus (30). In a study by the Baraiya Group in 2012, the application of
CMC in combination with clove essential oil effectively increased the shelf
life of tomato by delaying the ripening of the fruit and postponing their
senescence (31).


5. Conclusion

This study
showed that the CMC edible coating enriched with Zataria multiflora essential oil can maintain the fruit quality of fresh pistachios during storage.
Furthermore, this treatment prevents excessive weight loss and improves the
fruit quality in terms of phenolics, antioxidant activity, carbohydrates, and
TSS. We concluded that using an edible coating containing 1.5% (w/v) CMC enriched
with 0.4% (w/v) Zataria multiflora essential oil can extend the duration of storage and
maintain the quality of fresh pistachios for 32 days.